Legionellosis is a disease of significant medical and public interest. Legionella is commonly found in aquatic habitats where its ability to survive and to multiply within different protozoa equips the bacterium to be transmissible and pathogenic to humans. In addition Legionella has become a favored model system to analyse the mechanisms of bacterial survival, acquisition of nutrients and intracellular replication. Following the recent publication of the genome sequences of four L. pneumophila strains it is now feasible to investigate the whole genome in silico, the transcriptome via micro arrays and the proteome by two-dimensional gel electrophoresis. Research in the fields of clinical features, diagnosis, treatment and epidemiology continues to generate new data. The topics covered by this volume range from the history of the identification of Legionella and clinical disease treatment, to the microbe's gene expression and secretion systems as well as its strategies for intracellular multiplication and nutrient acquisition. The main focus of the book is the current state of many of the most critical features of Legionella. Internationally renowned authors have contributed chapters describing and discussing the latest research findings with an emphasis on molecular aspects. The editors and authors have produced an excellent book that will be an extremely useful reference source. This comprehensive publication is aimed at readers with teaching or research interests in microbiology, genetics, genomics, infectious diseases or clinical research. A recommended book for all microbiology and clinical research laboratories.

Reviews

"This book is a must for microbiologists involved with Legionella ... it will make a nice addition to the library of anyone professionally involved with Legionnaires' disease and Legionella bacteria."fromLegionella Enews (2008)

"This comprehensive review of the Legionella species covers the clinical presentation of disease in humans, treatment options, and epidemiology of
disease."fromDoodys (2008)

"The book is up to date and extensively referenced. The editors have sought contributions from renowned researchers in this field and the book would be a valuable source of information for scientists entering this field."fromACM News (2008) 2: 17

"an excellent reference book for scientists interested in the molecular biology of Legionella and its quality is attributed to the topical and interesting content, presentation and editorial style."fromAus. J. Med. Sci. (2009) 30: 106.

Table of contents

1. Legionnaires' Disease: History and Clinical Findings

Paul H. Edelstein

The history of Legionnaires' disease began at least 33 years before the 1976 Philadelphia epidemic, when Legionella micdadei was isolated from human blood. Multiple isolations of several different Legionella spp. were made prior to 1976, and it was known by 1968 that tetracycline therapy prevented deaths in L. pneumophila- infected chicken embryos. The 1976 epidemic provided the scientific focus and resources necessary to determine that L. pneumophila caused epidemic pneumonia and to show that epidemics of Legionnaires' disease had occurred worldwide many years before 1976. Despite a surfeit of available resources and expertise, the effort to isolate the etiologic agent succeeded solely on the basis of one person's determination to solve a scientific problem and his willingness to reexamine his assumptions about prior laboratory results. Pontiac fever, a disease of unknown etiology, is a self-limiting and short duration febrile illness that has been associated with exposure to L. pneumophila. Because of non-specific clinical findings that overlap with other diseases, accurate diagnosis of Pontiac fever in non-outbreak settings is impossible. Legionnaires' disease can be diagnosed specifically through specialized laboratory tests, but not by clinical findings alone. This is because the clinical findings of Legionnaires' disease overlap with those of other more common causes of community acquired pneumonia. Antimicrobial therapy of Legionnaires' disease requires the use of drugs that are active against intracellular Legionella spp., such as tetracyclines, macrolides, azalides and antibacterial quinolones.

2. Diagnostics and Clinical Disease Treatment

Paul C. Lück

The methods currently available to diagnose Legionnaires' disase are culture, urinary antigen detection, direct fluorescent antibody testing, detection of nucleic acid and detection of specific antibodies in serum samples. The advantages and limitations of each method are discussed. Presently, none of the diagnostic tests available offers the desired quality with respect to sensitivity and specificity. Culture should be obligatory, especially when hospitalized patients with underlying diseases are investigated. A positive culture is the prerequisite of molecular epidemiological investigations. Urinary antigen detection is a valuable tool in the majority of community-acquired cases when L. pneumophila serogroup 1 is the causative agent. In cases of nosocomial disease, when Legionella pneumophila serogroups other than sg 1 are frequent, this assay has limitations. The detection of nucleic acid is very promising, but needs further validation. The detection of antibodies in a patient's serum is of little use in the acute phase of the illness. Several molecular subtyping techniques are in use to subtype L. pneumophila strains in epidemiological investigations. Legionella pneumophila is genetically very heterogeneous thus allowing an individual fingerprint of each strain. However, the majority of clinical cases are caused by a limited number of clones that cause disease worldwide. The therapy of for Legionnaires' disease requires drugs that can access and are active intracellularly. Currently, fluorochinolones and macrolides are the most active agents.

3. The Epidemiology of Legionnaires' Disease

Carol Joseph and Katherine Ricketts

Much has been learnt about the epidemiology of Legionnaires' disease since the organism was first identified in 1976. National surveillance systems and research studies were established early, and in recent years improved ascertainment and changes in clinical methods of diagnosis have contributed to an upsurge in reported cases in many countries. Environmental studies continue to identify novel sources of infection, leading to regular revisions of guidelines and regulations. Between 1995 and 2005 over 32,000 cases of Legionnaires' disease and more than 600 outbreaks were reported to the European Working Group for Legionella Infections (EWGLI). In the future, there may be an increase in cases as the population becomes more elderly and climate change perhaps leads to an increased risk of exposure to infection in many countries. This would put ever more demands on public health resources for the management of this disease. This chapter on epidemiological aspects of Legionnaires' disease will discuss how surveillance, detection, treatment, control and prevention strategies, development and use of guidelines and legislation, and national and international public health initiatives all aim to improve our knowledge about the disease and contribute to reducing its public health burden on the population.

4. Developmental Cycle: Differentiation of Legionella pneumophila

Rafael A. Garduño, Audrey Chong and Gary Faulkner

The discovery that L. pneumophila follows a developmental cycle, within which it differentiates into numerous forms with highly altered phenotypes, underlines the usefulness of this bacterium as an experimental model for the study of bacterial intracellular differentiation. The developmental cycle of Chlamydia species (which are obligate intracellular pathogens) constitutes the current paradigm of bacterial intracellular differentiation. However, unlike Chlamydia, L. pneumophila can grow in vitro and is amenable to mutagenesis and broad genetic screenings, enhancing our investigative options. Both the extracellular and the intracellular developmental cycles of L. pneumophila have been described. The comparative analysis of the differentiation steps associated with these two developmental cycles suggests that the extracellular cycle is an incomplete derivative of the intracellular one. We propose that L. pneumophila possesses a single developmental program controlled by known regulators of bacterial differentiation that respond to changes in the levels of amino acids in the environment, and to other as yet unidentified intracellular factors. However, these regulators seem to play unique roles in L. pneumophila that deviate from their known functionality in other bacteria.

5. The Genetics and Immunology of Host Resistance to Legionella Infection

Russell Vance and Thomas R. Hawn

Significant progress has been made toward elucidating the critical genetic and immunological components of the host immune response to Legionella pneumophila. In this chapter, we review the host immune response to Legionella, including the roles of the innate immune response, B cells, T cells, cytokines and chemokines. We examine the use of in vitro studies of macrophage infection, in vivo animal studies, and more recent human genetic studies that have all contributed to our understanding of the immune response to Legionella. We also highlight the distinct biological properties of Legionella that have stimulated discoveries of broad relevance to our understanding of host-pathogen interactions. Taken together, numerous genetic and immunological studies have revealed that Legionella is a fascinating model for understanding how the host interprets a large number of molecular and cellular cues to provide host defense.

6. The Flagellar Regulon of Legionella pneumophila and the Expression of Virulence Traits

Klaus Heuner and Christiane Albert-Weissenberger

Legionella is a ubiquitious inhabitant of aquatic habitats, and it is believed that motility is an important feature in the life cycle of L. pneumophila in its environment. To survive, L. pneumophila must be able to respond to different environmental factors and to modulate gene expression. Furthermore, very early after the first report of Legionnaires' disease, researchers discovered that most Legionella strains are flagellated and that regulation of the flagellum and the expression of virulence traits are linked. Expression of the flagellum is modulated by various environmental factors. The flagellar regulon, comprised of genes encoding the structural components of the flagellum, is controlled by a cascade of regulators. This chapter will focus on the composition of the flagellar complex and the regulation of flagellar operons, including the virulence traits of L. pneumophila that are coordinately expressed with the flagellar genes. The contribution of the flagellar system to the virulence of L. pneumophila will also be discussed.

7. Genomics and Transcriptomics of Legionella pneumophila: Insights into the Life Style of an Intracellular Pathogen

Christel Cazalet, Matthieu Jules and Carmen Buchrieser

The last years have seen a giant step in genomics of Legionella pneumophila. The determination and publication of the complete genome sequences of three clinical L. pneumophila isolates in 2004 now paves the way for major breakthroughs in understanding the biology of L. pneumophila in particular and Legionella in general. Here we describe the major features deduced from genome sequence analyses of L. pneumophila and highlight the characteristic features and common traits of the L. pneumophila genomes. Emphasis is given to putative virulence and Legionella life cycle related functions. In depth comparative genome analysis using DNA arrays to study the gene content of 180 Legionella strains revealed the high genome plasticity and frequent horizontal gene transfer identified through whole genome comparisons. Further insight in the L. pneumophila life cycle was recently gained by investigating the intracellular gene expression profile of L. pneumophila in Acanthamoeba castellanii, its natural host. This study shows that L. pneumophila exhibits a biphasic life cycle in vitro as well as in vivo and defines transmissive and replicative traits according to gene expression profiles. These specific traits are discussed, and future perspectives in Legionella genomics are presented.

8. Secretion and Export in Legionella

Nicholas P. Cianciotto

L. pneumophila secretes many factors that promote its growth and persistence within the environment, various types of host cells, and the mammalian lung. These factors include both proteins and non-proteinaceous molecules. This chapter will focus on two topics. First will be the type II protein secretion system that elaborates a large number and wide variety of enzymes, some of which have recently been implicated as factors that promote infection. Second will be the export of siderophores, non-protein, high-affinity iron chelators that stimulate growth in low-iron conditions.

9. The Dot/Icm Type IVB Secretion System of Legionella

Carr D. Vincent and Joseph P. Vogel

The major virulence system of Legionella pneumophila is a specialized secretion system encoded by twenty-six dot/icm genes. This secretion machine is classified as a type IVB secretion system due to its homology to the transfer apparatus of IncI conjugative plasmids. In Legionella, the Dot/Icm secretion system is used to export a large number of protein substrates into the cytoplasm of phagocytic cells, thereby allowing Legionella to survive and replicate within these normally bacteriocidal cells.

10. Mechanisms of Intracellular Survival and Replication of Legionella pneumophila

Tamara O'Conner, Matthew Heidtman and Ralph R. Isberg

Upon uptake by phagocytic cells the Legionella containing phagosome (LCV) evades targeting to and degradation by the endocytic pathway. Instead, mitochondria and early secretory vesicles are rapidly recruited to the LCV. Over a period of several hours, the LCV becomes studded with ribosomes and resembles an ER-like compartment. Within this organelle L. pneumophila is able to replicate to high numbers. Intracellular growth of L. pneumophila is dependent on the dot/icm genes. Many of the Dot/Icm proteins assemble into a large membrane-spanning complex that enables the delivery of bacterial proteins into the host cell cytosol. These translocated substrates are thought to modulate a variety of host cell processes to create an intracellular environment permissive for bacterial replication. The functions of most of these substrates are unknown, although at least some of them are known to modulate host cell vesicle trafficking and anti-apoptotic signaling pathways. Recent studies have sought to identify host cell processes important for L. pneumophila intracellular replication. The input of multiple host cell vesicle trafficking pathways, the ERAD system, anti-apoptotic signaling, autophagy, and host cell lipid metabolism all appear to be important to varying extents for efficient intracellular growth of L. pneumophila.

The fitness of the opportunistic pathogen L. pneumophila is a result of selective pressures in its natural freshwater environments. L. pneumophila has thus developed a sophisticated morphogenetic code that equips it to dramatically alter its developmental state in response changes in its microenvironment. It is now clear that the cellular and molecular transformations that occur during the L. pneumophila life cycle are integrated with its metabolic state. This is exemplified by its reciprocal expression of traits that promote replication within host cells and those that promote transmission to a new host or its differentiation into cyst-like forms of greatly reduced metabolic activity. Insight into some aspects of the metabolic machinery that equip L. pneumophila to satisfy its nutritional requirements have been gained through studies of the molecular mechanisms that promote its differentiation. Availability of the genome sequences of L. pneumophila strains Philadelphia-1, Paris and Lens has also increased our power to decipher the strategies L. pneumophila employs to acquire and assimilate nutrients from its environment. By integrating knowledge gained from genetic, biochemical, molecular and computational approaches, this chapter presents our current view of the metabolic capabilities of L. pneumophila, focusing on its key metabolic features, nutritional requirements and specialized nutrient acquisition systems.

12. Dictyostelium, a Tractable Model Host Organism for Legionella

Heike Bruhn and Michael Steinert

The haploid amoeba Dictyostelium has proven to be a suitable model system for studying cellular aspects of Legionnaires' disease. In this review we describe both Legionella and Dictyostelium factors and processes that are relevant to infection. Moreover, we summarize the results of the Dictyostelium transcriptional host cell response to infection and discuss how genetic and molecular biology techiques available for Dictyostelium can further improve our knowledge of Legionella-host cross-talk.